The detection of obligate anaerobic bacteria in swine abscesses. A comparison between gas-liquid chromatography and bacteriological culturing methods

From the 62 swine abscesses examined, 56 yielded positive cultures, of which 38 cases were mixtures of facultative aerobes and obligate anaerobic bacteria (OAB). Only 6 species appeared to be sterile. In all specimens, which yielded OAB, VFA (C4-C6) could be detected by gas-liquid chromatography (GLC). False negatives were not recorded, and only one false positive was found, which might have been due to faulty culture technique, or to the fact that the metabolic markers may persist longer than the OAB themselves. Because current methods to isolate non-sporulating OAB are laborious and not available in most veterinary clinical situations and because there is a need to evaluate the importance of non-sporulating OAB in veterinary medicine, GLC seems to be a useful alternative to culture techniques for detecting the presence of OAB in infectious processes in animals.     

Summer mastitis in heifers: a bacteriological examination of secretions from clinical cases of summer mastitis in Denmark

A total of 166 samples of secretions collected from Danish heifers suffering from clinically diagnosed summer mastitis were examined bacteriologically. One hundred and sixty three samples yielded positive findings whereas no growth was obtained from 3 specimens. The majority of samples revealed a mixed flora in which the predominant components were Actinomyces pyogenes, Peptostreptococcus indolicus, a microaerophilic coccus (Stuart-Schwan coccus), Fusobacterium necrophorum, Bacteriodes melaninogenicus and Streptococcus dysgalactiae. Pure cultures were recovered in only 7% of cases. P. indolicus was isolated from 875 of the cases, a microaerophilic coccus from 84%, A. pyogenes from 72%, Fusobacterium necrophorum from 52%, Str. dysgalactiae from 37%, B. melaninogenicus from 33% and various unidentified obligate anaerobic Gram-negative rods from 31%. The results confirm the complex bacterial aetiology of summer mastitis.     

Comparative bacteriological studies on summer mastitis in grazing cattle and pyogenes mastitis in stabled cattle in Denmark.

A total of 143 secretions from clinical cases of summer mastitis (SM) in grazing cattle and from 89 cases of pyogenes mastitis (PM) in stabled cattle were examined bacteriologically. The typical bacteriological finding was a mixed flora in which the predominant organisms were Actinomyces pyogenes (SM-70%, PM-85%), Peptostreptococcus indolicus (54%, 54%), a microaerophilic coccus (Stuart-Schwan coccus) (26%, 25%), Fusobacterium necrophorum biovar B (22%, 12%), Bacteroides melaninogenicus (20%, 9%) and Streptococcus dysgalactiae (21%, 5%). All except six cases occurred in non-lactating animals or within three weeks after parturition. The majority of animals (about 90%) had only one quarter affected and no differences in quarter distribution were observed between the two groups.     

Rapid presumptive diagnosis of anaerobic infections in animals by gas-liquid chromatography

The detection of volatile fatty acids (VFA) by gas chromatography of 85 purulent specimens from abscesses or pyogenic infections in cats, dogs, rodents, and ruminants was compared with the results of bacteriologic culturing, and proved to be a rapid means of presumptively diagnosing anaerobic infections. Of 83 bacteriologically positive specimens, 52 (61%) yielded obligate anaerobes and in 50 specimens, 1 or more VFA (butyric acid, isobutyric acid, valeric acid, isovaleric acid, caproic acid, or isocaproic acid) was detected. Forty-six specimens were positive for culturing of anaerobes and for detection of 1 or more of these VFA. By contrast, pus from infections caused by (facultative) aerobic microorganisms contained no VFA or only acetic and/or propionic acid.     

A mild outbreak of bovine mastitis associated with Mycoplasma bovigenitalium.

Mycoplasma bovigenitalium was isolated from milk samples from 16 of 99 cows on one farm during a 15-week period in the summer of 1986. One cow was severely affected, four cows had relatively mild signs of mastitis, and three had only altered dry-cow udder secretions. Eleven of the infected cows were dry and three had been calved less than 48 hours. The abrupt method of drying-off and improvements in cleaning of the milking equipment were introduced, but no other control measures were instituted to eradicate the mycoplasma infections. After this mild outbreak of mastitis the herd was monitored for the next 17 months. In total 19 cows had a mycoplasma isolated from udder secretions. Acholeplasmas were isolated from 14 cows but were not associated with clinical mastitis. The udder infections with mycoplasmas apparently resolved without resorting to the segregation and culling of infected animals.     

Clinical, bacteriological and therapeutic aspects of bovine mastitis caused by aerobic and anaerobic pathogens.

Twenty cows and three heifers with summer mastitis-like signs were included in the study. Forty aerobic and 38 anaerobic bacterial isolates representing 23 different species were isolated from udder secretions, the mean number of species being 3.4/sample. Pathogens most often were recovered in quantities exceeding 10(5) CFU/ml. Growth of pathogenic bacteria in the blood was found in only one case. Most cases occurred in housed animals and had often been preceded by teat injury. No anaerobic growth was found in milk samples of a control group consisting of 29 cows with ordinary acute mastitis. Values of indicators of inflammation in milk (NAGase and plasmin activity) were high, mean values being 914.1 and 1.3 units. The animals with aerobic/anaerobic mastitis were treated using various antimicrobial agents. All but three quarters were lost to milk production. Two animals with complete recovery were given a combination of penicillin G and tinidazole.     

Mastitis in lactating camels (Camelus dromedarius) in Afar Region, north-eastern Ethiopia.

Quarter milk samples (n = 543) from 152 traditionally managed lactating camels (Camelus dromedarius) in Afar Region, north-eastern Ethiopia were examined to determine the prevalence of camel mastitis and identify its bacterial causes. Out of 152 camels examined, 19 (12.5%) were diagnosed as clinical mastitis cases based on clinical signs and bacteriological examinations. Of the 257 California Mastitis Test (CMT) positive quarter milk samples 162 (63.0%) yielded pathogenic bacteria. A positive correlation was observed between CMT positive results and presence of major pathogens in camel milk samples. The main mastitis pathogens isolated were Staphylococcus aureus, coagulase-negative staphylococci, Streptococcus agalactiae, S. dysgalactiae, and other species of streptococci, Pasteurella haemolytica and E. coli. Results of the present study suggest that mastitis in Afar camels is prevalent, Gram-positive cocci are the major isolates from camel milk samples and the CMT can be used as a screening test for the detection of mastitis in camels.     

Enzootic mycoplasmal mastitis in a large dairy during an eight-year period

During an 8-year period, a study was conducted in a large dairy in the Sacramento Valley of California, to define factors associated with the introduction and spread of mycoplasmal mastitis. To identify cows in which mycoplasmal infection appeared in 22 of 25 (88%) periods of new infection, milk samples were collected weekly from freshened cows and from hospitalized cows with clinical mastitis. The disease first appeared in freshened cows in at least 36% of the periods of new cases of mycoplasmal mastitis, which indicated that special attention must be paid to freshening cows in an attempt to control spread of the disease. New cases of mycoplasmal mastitis were recorded more often in January through April than during the rest of the year. Rates of mastitis infection caused by other pathogens (contagious and environmental organisms) increased during months when new cases of mycoplasmal mastitis were recorded. Sanitation on the farm during udder infusion and milking was a major factor in controlling the introduction and spread of mycoplasmal mastitis.     

The occurrence of summer mastitis in Jutland (Denmark) in relation to meteorological factors

Data for incidence of summer mastitis in 14 veterinary practices in Jutland (Denmark) were related to meteorological factors using statistical methods. A correlation and regression analysis showed that summer mastitis occurred earlier in years with high temperature and precipitation in early summer. The annual number of cases markedly increased over the years 1953 to 1980. This increase was described by applying a linear increase with with year to a logarithmic transformation of the annual incidence.Analysis of 10-day incidences from the veterinary practices and daily incidence from a test site at St. Vildmose (Jutland) showed that incidence of summer mastitis could be related to temperature and wind velocity within the previous 10 days. High temperature and low wind velocity in this period increase the incidence. This indicates that the incubation period of summer mastitis is at maximum about 10 days. The activity of the fly (Hydrotaea irritans Fall), which acts as mechanical vector for the bacteria is influenced by temperature and wind velocity. This may explain the variation in incidence of summer mastitis during the season and from year to year.     

Limulus test--a rapid and simple method for the detection of endotoxins produced by gram-negative bacteria in mastitis milk

According to the present study the limulus amebocyte lysate test (LAL) seems to be a convenient test to detect endotoxin in milk from udder quarters with and without inflammation. The correlation between endotoxin concentration and the results from the bacteriological investigation of 79 milk samples was good (Table I). Determination of endotoxin in 20 milk samples from cases of acute clinical mastitis with high cell count and a negative bacteriological culture showed that all but one had an endotoxin concentration of greater than 1.0 ng/ml milk (Table II). By using a micromethod of the LAL it is possible to detect cases of mastitis caused by gram-negative bacteria about one hour after the sample has reached the laboratory. In a preliminary field study milk from 13 cases of acute clinical mastitis were tested by a modified limulus amebocyte lysate (LAL) test ("cowshed test"). A 100% correlation to bacteriological findings was observed (Table IV). By using the LAL test to detect mastitis cases caused by gram-negative bacteria great economic advantages and less risk for resistance problems can be achieved by using proper antibiotics. This is the fact in Sweden where the frequency of acute clinical mastitis caused by streptococci (100% of strains sensitive for penicillin) and Staphylococcus aureus (about 90% of strains sensitive for penicillin) is high (70-80%) and about 20% are caused by gram-negative bacteria, mostly E. coli.     

Effect of sampling method and incubation temperature on fungal culture in canine sinonasal aspergillosis

O bjectives : To evaluate the most appropriate sampling procedure and the effect of incubation temperature on fungal culture in the diagnosis of canine sinonasal aspergillosis (SNA).
M ethods : Sixteen dogs with SNA and 20 dogs with non-fungal nasal disease entered a prospective study. Nasal secretions and mucosal biopsies were collected in all dogs. Fungal plaques were also sampled in dogs with SNA. Each specimen was taken in duplicate from each dog and incubated at room temperature and 37°C.
R esults : In dogs with SNA, nasal secretions, mucosal biopsies and fungal plaques yielded fungal growth at room temperature in one, one and seven dogs, respectively, whereas fungal growth was obtained at 37°C in three, 12 and 14 dogs, respectively. No specimen collected from any dog with non-fungal nasal disease yielded fungal growth at room temperature or at 37°C.
C linical S ignificance : The diagnosis of canine SNA is more likely to be confirmed following culture of mucosal biopsies or fungal plaques than nasal secretions sampled blindly with swabs. Incubating cultures at 37°C is more likely to provide a diagnostic outcome than when samples are cultured at room temperature. Fungal culture of nasal specimens has good specificity for the diagnosis of SNA in dogs.     

Occurrence of clinical and sub-clinical mastitis in dairy herds in the West Littoral Region in Uruguay

Twenty-nine dairy farms were selected to determine the incidence of clinical mastitis, prevalence of sub-clinical mastitis and bacterial aetiology in the West Littoral Region of Uruguay. In samples taken by the owner and frozen at -20 degrees C during a week the incidence rate of clinical mastitis was determined as 1.2 cases per 100 cow-months at risk. Staphylococcus aureus was the most common isolated pathogen in 37.5% of 40 milk samples from clinical cases obtained in 1 month. No bacteria grew in the 32.5% of the total samples. A sub-sample including 1077 dairy cows from randomly selected farms was used to determine the prevalence of sub-clinical mastitis. These samples were taken on one visit to each farm. The prevalence was 52.4% on a cow basis and 26.7% on an udder quarter basis. In 55.1% of the quarters of the selected animals with more than 300,000 cells/ml there was no growth. The isolated pathogens from sub-clinical cases and their relative frequencies were: Staphylococcus aureus 62.8%, Streptococcus agalactiae 11.3%, Enterococcus sp. 8%, coagulase-negative staphylococci 7.4%, Streptococus uberis 6.4%, Streptococcus dysgalactiae 1.8%, Escherichia coli 1.5% and Staphylococcus hyicus coagulase-positive 0.6%.     

Cause, occurrence, and clinical signs of mastitis and anorexia in cows in a Wisconsin study

Microbiologic culture revealed the following cause of mastitis and anorexia in 145 cows in Wisconsin to be Escherichia coli, 66 cows; Klebsiella spp, 3; Corynebacterium pyogenes, 27; streptococci, 21; staphylococci, 20; yeasts, 1; and no bacterial growth, 7. Mastitis was detected with approximately equal frequency throughout the year. Escherichia coli was isolated throughout the year, but was more common and was the predominant organism during the summer. Corynebacterium pyogenes was isolated most often in winter and spring; streptococci in fall, winter, and spring; and staphylococci throughout the year. Corynebacterium pyogenes caused most of the mastitis in nonlactating cows. Escherichia coli, C pyogenes, streptococci, and staphylococci were isolated with about equal frequency at parturition, whereas E coli was the predominant cause of mastitis in early and late lactation. Of cases of mastitis, 27% were seen 10 days before and after parturition. Local and systemic clinical signs of infection were similar for all causes, except that C pyogenes caused more (P less than 0.01) malodorous and purulent milk than did other organisms and was isolated more commonly from quarters with injured teats. Recovery was significantly (P less than 0.01) higher in cows with E coli infections, compared with recovery in cows with gram-positive organism infections. Cows with C pyogenes infections frequently had quarters that ultimately ceased lactation. A few cows were recumbent at initiation of antimicrobial therapy and a few were not eating 24 hours later; however, 50% of these cows recovered. Criteria such as season of year, stage of lactation, appearance of milk and udder, and appetite permitted the cause (gram-negative or gram-positive organisms) of the mastitis to be predicted with 77% accuracy.     

Ruminal evacuation's effect on microbial activity and ruminal function

The influence of evacuating, mixing and returning ruminal contents on microbial populations, volatile fatty acid (VFA) concentrations and liquid flow rate was investigated with four ruminal-cannulated Hereford steers (247 kg avg wt). Ruminoreticular contents were sampled, then completely removed, mixed for 5 min and returned to the rumen. Subsequent samples were taken immediately, 1 h and 4 h later. Non-evacuated steers were sampled at identical time intervals either 1 d before or after evacuation. Averaged over time, there was no significant difference between evacuated and non-evacuated steers in total anaerobic, cellulolytic and facultative bacteria, protozoa, oxidation-reduction potential, VFA concentrations, and liquid flow rates. There were no treatment X time interactions and, except for holotrich protozoa and VFA, no differences from time of sampling. Ruminal evacuation does not appear disruptive to anaerobiosis or detrimental to ruminal microorganisms and digestive processes.     

Sources of Prototheca spp in a dairy herd environment

A source of Prototheca spp causing mastitis in a herd of 263 milking cows in North Carolina was investigated. Of 38 samples from the dairy environment, 18 (47%) contained Prototheca spp. Isolation sites included cattle drinking water; a feed trough; mud, dirt, and excreted feces from a dirt lounging area; water, sludge, mud, and vegetation from a creek in the lounging area; and the floor of a freestall barn. Samples were collected from 5 additional dairies, including one dairy with and 4 dairies without a history of protothecal mastitis. Prototheca spp were isolated from 48 (25%) of 190 samples from various sites on all 6 dairies. Isolates were P zopfii (45; 94%) and P wickerhamii (3; 6%). Isolation frequency ranged from 4 to 47% of samples/dairy. There was no apparent difference in the isolation frequency of Prototheca spp from samples from dairies with or without a history of protothecal mastitis. Sites characterized by wetness and the presence of organic matter most commonly yielded Prototheca spp. Because Prototheca spp appear to be common in the dairy environment, factors in addition to presence in the environment may be important in development of protothecal mastitis.     

Bacteria associated with pyothorax of dogs and cats: 98 cases (1989-1998)

OBJECTIVE: To determine the organisms most commonly isolated from pleural fluid from dogs and cats with pyothorax. DESIGN: Retrospective study. ANIMALS: 51 dogs and 47 cats. PROCEDURE: Results of bacteriologic culture of pleural fluid samples obtained by means of thoracentesis were obtained from medical records. To obtain information on in vitro antimicrobial susceptibility of organisms commonly isolated from dogs and cats, records of all dogs and cats examined during 1998 were reviewed, and information was obtained on identity and in vitro antimicrobial susceptibility of aerobic organisms isolated from samples other than urine or urinary tract samples. RESULTS: Median ages of dogs and cats were 4 years. Bacteria were isolated from pleural fluid samples from 47 of 51 (92%) dogs and 45 of 47 (96%) cats. Obligate anaerobic bacteria were isolated from 28 dogs and 40 cats. A mixture of obligate anaerobic and facultative bacteria was isolated from 17 dogs and 20 cats. Samples from cats most often yielded a member of the nonenteric group (most commonly members of the genus Pasteurella), whereas those from dogs more often yielded a member of the family Enterobacteriaceae (most commonly E coli). CONCLUSIONS AND CLINICAL RELEVANCE: Results suggest that antimicrobial agents chosen for the initial treatment of dogs and cats with pyothorax should be active against a mixture of obligate anaerobic and facultative bacteria.     

The role of intestinal volatile fatty acids in the Salmonella shedding of pigs

Factors affecting colonization of the intestinal tract by salmonellas were studied in two pig herds. In herd H 18% of the faecal samples taken from live pigs and 30% of the colon content samples collected at slaughter contained salmonellas. In contrast, the 50 faecal samples taken from pigs of herd L were negative and only 2% of the colon contents collected at the yielded salmonellas. An antibacterial effect inhibiting salmonella multiplication was demonstrable in vitro in colon contents from pigs of herd L. No such effect was found to exist in samples taken from pigs of herd H. The antibacterial effect is due to the non-dissociated volatile fatty acid (VFA) molecules present in the colon content. As the degree of VFA dissociation depends on the pH of the environment, at lower pH values (pH 6.1 +/- 0.2) of the colon contents from herd L the ratio of non-dissociated VFA molecules is higher and the resulting antibacterial effect is stronger than in samples from herd H (pH 7.1 +/- 0.3).     

Concentration of volatile fatty acids in digesta samples obtained from healthy cows and cows with cecal dilatation or dislocation

OBJECTIVE: To compare concentrations of acetic, propionic, butyric, and i- and n-valerianic acids in digesta samples obtained from the rumen, cecum, proximal loop of the ascending colon (PLAC), and rectum of healthy cows and cows with cecal dilatation or dislocation (CDD). ANIMALS: 20 cows with CDD and 20 healthy cows. PROCEDURE: Samples were collected from all sites during surgical correction of CDD and also from the rectum 1, 2, and 3 days after surgery (group CDD). Samples from healthy (control) cows, matched on the basis of diet and milk yield, were obtained at a slaughterhouse. Concentrations of volatile fatty acids (VFA) were analyzed by use of gas chromatography. Absolute concentration of each VFA was additionally corrected for pH to allow calculation of the concentration of undissociated VFA. RESULTS: Absolute concentration and concentration of the undissociated form of all analyzed VFA were significantly increased in samples collected from the cecum and PLAC of cows in group CDD, compared with concentrations for control cows. Within 3 days after surgery, significant decreases of the absolute concentration of butyric, i- and n-valerianic acids, and undissociated i- and n-valerianic acids were evident in samples obtained from the rectum of group-CDD cows. Concentrations of VFA in samples obtained from the rectum during surgery correlated with corresponding VFA concentrations in samples obtained from the PLAC. CONCLUSIONS: Concentrations of VFA are increased in the cecum and PLAC of cows with CDD. However, the role of increased concentrations of VFA in the etiopathogenesis of CDD is unknown.     

Prevalence of mycoplasmal bovine mastitis in California

Seven species of mycoplasma plus one or more unknown species were found to cause bovine mastitis in California. Both the frequency of cases and number of species of mycoplasma in samples received at the laboratory have increased from 1976 to 1978. By survey, nearly 4% of samples of bulk tank milk from dairy farms were found to contain mycoplasma of potential pathogenic significance. Acholeplasma laidlawii was frequently isolated from samples both from cows and from farm bulk tanks during wet, rainy weather in the spring of 1978, apparently as contaminants only. The prevalence of positive bulk tank milk samples in an area appeared to parallel the prevalence of clinical mycoplasmal mastitis problem herds.     

奶牛隐性乳房炎奶样中埃希氏大肠杆菌共同性抗原初探

用LMT的方法对新疆8个奶牛场采集的1020份奶样进行隐性乳房炎的检测,并对阳性样品进行了大肠杆菌的分离与鉴定。从不同牛场分离菌株中挑选8株用家兔制备免疫血清,用试管凝集对从乳房炎奶样分离出的其它21株大肠杆菌进行共同性抗原的筛选。结果显示,免疫血清效价达1∶2560～1∶5120,85.7%的待检菌株与其中3种血清凝集反应呈阳性。对制备血清的8株菌进行SDS-PAGE电泳分析,菌体蛋白图谱显示在20~84 ku处存在大量相同或相似的蛋白条带;用制备的免疫血清进行免疫印迹分析,与8株菌的菌体蛋白在70 ku处呈现明显的抗原抗体反应。说明这些菌株存在共同性抗原。